Positron Emission Tomography (PET) is a molecular imaging technology that is used effectively for the detection of disease. PET imaging systems create images based on the distribution of positron-emitting isotopes in the tissue of a patient. The isotopes are typically administered to a patient by injection of probe molecules that comprise a positron-emitting isotope, such as F-18, C-11, N-13 or O-15, covalently attached to a molecule that is readily metabolized or localized in cells (e.g., glucose) or that chemically binds to receptor sites within cells. In some cases, the isotope is administered to the patient as an ionic solution or by inhalation. One of the most widely used positron-emitter labeled PET molecular imaging probes is 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG).
PET scanning using the glucose analog [18F]FDG, which primarily targets glucose transporters and hexokinase, is an accurate clinical tool for the early detection, staging, and restaging of cancer. PET-FDG imaging is also used to monitor cancer chemotherapy and chemoradiotherapy because early changes in glucose utilization have been shown to correlate with outcome predictions. A characteristic feature of tumor cells is their accelerated glycolysis rate, which results from the high metabolic demands of rapidly proliferating tumor tissue. Like glucose, FDG is taken up by cancer cells via glucose transporters and is phosphorylated by hexokinase to FDG-6 phosphate. FDG-6 phosphate cannot proceed any further in the glycolysis chain, or leave the cell due to its charge, allowing cells with high glycolysis rates to be detected.
Although useful in many contexts, limitations of FDG-PET imaging for monitoring cancer exist as well. Accumulation in inflammatory tissue limits the specificity of FDG-PET. Conversely, nonspecific FDG uptake may also limit the sensitivity of PET for tumor response prediction. Therapy induced cellular stress reactions have been shown to cause a temporary increase in FDG-uptake in tumor cell lines treated by radiotherapy and chemotherapeutic drugs. Further, physiologically high normal background activity (e.g. in the brain) can render the quantification of cancer-related FDG-uptake impossible in some areas of the body.
Due to these limitations, other PET imaging tracers are being developed to target other enzyme-mediated transformations in cancer tissue, such as 3′-[F-18]Fluoro-3′-deoxythymidine (FLT) for DNA replication, and [C-11](methyl)choline for choline kinase, as well as ultra high-specific activity receptor-ligand binding (e.g., 16α, [F-18]fluoroestradiol) and potentially gene expression (e.g., [F-18]fluoro-ganciclovir). Molecularly targeted agents have demonstrated great potential value for non-invasive PET imaging in cancers.
These studies have demonstrated the great value of non-invasive PET imaging for specific metabolic targets of cancer. Despite the clear clinical value of incorporating PET imaging into patient management, limitations do exist. In certain instances, current imaging probes lack specificity or have inadequate signal to background characteristics. In addition, new biological targets that are being tested for therapeutic intervention will require new imaging probes to evaluate therapeutic potential.
Thus, additional biomarkers are needed that show a very high affinity to, and specificity for, tumor targets to support cancer drug development and to provide health care providers with a means to accurately diagnose diseases and monitor treatment. Such imaging probes could dramatically improve the patient's outcome, allowing smaller tumors to be detected, with only nanomole quantities of the tracer injected into patients.
Key to the clinical success of cancer treatment is the ability to predict how a particular type of cancer will respond to treatment. In the specific case of tumors, factors such as the tumor's phenotype, size and location all dramatically affect therapeutic treatment decisions. While standard chemotherapeutic or radiation regimens are employed to treat a variety of tumors, certain tumor types resist standard therapeutic regimens and thus may worsen a patient's clinical outcome.
Due to the unique nature of cancer cell growth, its proliferative nature can offer clues for its therapeutic treatment. For instance, because of the rapid and disorganized growth of cancerous tumors, they oftentimes develop disorganized neovascularization leading to poorly vascularized environments. (Wang, J. and L. Maurer, Positron Emission Tomography: Applications in Drug Discovery and Drug Development Curr. Top. Med. Chem., 2005, 5: p. 1053-1075). In turn, environments that are removed 100-200 μm from blood supplies can become hypoxic, characterized by a tissue pO2 of less than 10 mmHg. In response to these hypoxic conditions, tumor overexpression of hypoxia inducible factor-1 (HIF-1) leads to the up-regulation of several proteins necessary for tumor survival including vascular endothelial growth factor (VEGF), carbonic anhydrase-IX (CA-IX), and glycolysis enzymes.
Hypoxic tumors are clinically problematic: they resist both the effects of radiation and cytotoxic therapy which can result in treatment failure. (Adams, G., Hypoxia-mediated drugs for radiation and chemotherapy. Cancer, 1981. 48: p. 696-707; Moulder, J. and S. Rockwell, Hypoxic fractions of solid tumors: experimental techniques, methods of analysis, and a survey of existing data. Int. J. Radiat. Oncol. Biol. Phys., 1984, 10: p. 695-712; Nordsmark, M., M. Overgaard, and J. Overgaard, Pretreatment oxygenation predicts radiation response in advanced squamous cell carcinoma of the head and neck. Radiother. Oncol, 1996. 41: p. 31-39.) Moreover, hypoxic cancer cells have been linked to malignant cancers that are known to spread invasively throughout the patient. (Brizel, D. M., et al., Tumour oxygenation predicts for the likelihood of distant metastases in human soft tissue sarcoma. Cancer Res., 1996. 56: p. 941-943; Höckel, M., et al., Association between tumor hypoxia and malignant progression in advanced cancer of the cervix. Cancer Res., 1996. 56: p. 941-943.) Several types of human cancers are well known to become hypoxic including breast, cervical cancer and non-small cell lung cancer. (Vaupel, P., et al., Oxygenation of human tumours: evaluation of tissue oxygen distribution in breast cancers by computerized O2 tension measurements. Cancer Res., 1991, 51: p. 3316-3322.)
Because hypoxic tumors respond poorly to both traditional radiation and cytotoxic therapies, several alternate approaches exist for treating hypoxic cancer cells including hyperbaric O2, ARCON, radiosensitizers and bioreductive cytotoxic agents. (Seddon, B. M. and P. Workman, The role of functional and molecular imaging in cancer drug discovery and development Brit, J. Radial., 2003. 76: p. S 1 28-S 138.) In the last example, the bioreductive agents containing the nitroimidazole chemotype are reduced intracellularly, forming radical anion metabolites that eventually become trapped intracellularly. In oxic environments, the radical anion reacts with O2 and returns to its premetabolized state.
Confirmation of tumor hypoxia in patients is necessary in order to appropriately plan bioreductive-based therapies for treatment. Determining tumor hypoxia via electrode measurements of pO2 concentrations within the tumor is an impractical endeavor at best. In addition, it is only possible to interrogate superficial tumors with this technique. A more general and less invasive method for detecting the hypoxic nature of cancer cells relies on radioactively labeled, bioreducible tracers that localize within hypoxic cells inversely proportional to their cellular pO2.
There are several bioreducible imaging agents that can detect hypoxic cells in vivo including [18F]F-MISO (Rasey, J. S., et al., Determining the hypoxic fraction in a rat glioma by uptake of radiolabeled fluoromisonidazole. Radiat. Res., 2000. 153: p. 84-92; Bentzen, L., et al., Feasibility of detecting hypoxia in experimental mouse tumours with 18F-fluorinated tracers and positron emission tomography: a study evaluating 18F-Fluoromisonidazole and [18F]Fluoro-2-deoxy-D-glucose. Acta. Oncol., 2000. 39: p. 629-637), [18F]F-EF1 (Hustinx, R., et al., Non-invasive assessment of tumor hypoxia with the 2-nitroimidazole 18F-EFI and PET J. Nucl. Med., 1999. 4: p. 99P (abstract 401)), [18F]-FETNIM (Chao, K. S., et al., A novel approach to overcome hypoxic tumor resistance: Cu-ATSM-guided intensity-modulated radiation therapy. Int. J. Radiat. Oncol. Biol. Phys., 2001. 49: p. 1171-1182; Yang, D. J., et al., Development of 18F-labeled fluoroerythronitroimidazole as a PET agent for imaging tumor hypoxia. Radiology, 1995. 194: p. 795-800; Gronroos, T., et al., Pharmacokinetics of [18F]-FETNIM. A potential hypoxia marker for PET J. Nucl. Med., 2001. 42: p. 1397-1404), [18F]FRP-170 (Ishikawa, Y., et al., Development of [18F]FRP-170 injection for imaging hypoxia by PET., Kaku Igaku., 2005, 42: p. 1-10.), and [62Cu]-ATSM (Fujibayashi, Y., et al., Copper-62-ATSM—a new hypoxia imaging agent with high membrane permeability and low redox potential. J. Nucl. Med., 1997. 38: p. 1155-1160).
One of most clinically studied hypoxia markers is [18F]F-MISO, the fluorine analog of the hypoxic cell radiosensitizer misonidazole. [18F]F-MISO successfully identifies hypoxic tumors in patients, however, its slow diffusion into hypoxic tumors requires longer uptake times before imaging and, in addition, high background uptake leads to small tumor to background ratios. As an alternative, several groups have prepared nitroimidazoles with less lipophilic character in an attempt to increase the tumor to background ratio by increasing the tracer's washout from normoxic tissue.
While these hypoxic imaging agents show clinical promise, there exists an unmet need for hypoxia tracers that possess enhanced pharmacokinetic profiles leading to peak signal to noise ratios in shorter periods for faster and potentially more accurate hypoxia assessments.